Cells are composed of thousands of different molecules. To study
these molecules, scientists must be able to isolate them in pure form. To
isolate just one type of biomolecule from the thousands of others present in a
cell can be a daunting task, like looking for a needle in haystack. Fortunately,
powerful separation techniques developed over the past century make it
possible to isolate virtually every type of molecule known to occur in living
cells. Two of the most powerful techniques for purifying biomolecules are
chromatography and electrophoresis. In this lab you will use chromatography to
separate a mixture of lipid-soluble pigments extracted from spinach leaf cells.
In labs 8 and 9 you will use electrophoresis to separate
mixtures of proteins and nucleic acids.

Use the interactive
exercise below to learn more about chromatography.

The pigments present in spinach leaf cells are used to absorb light energy needed for
photosynthesis. The color of
each pigment depends on which wavelengths of visible light it reflects and which
wavelengths it absorbs. Green pigments, for example, reflect most of the
wavelengths in the green region while they strongly absorb light in the red and
blue regions. A
scanning spectrophotometer is an instrument that can automatically measure how much light at
each wavelength is absorbed by a given pigment, over an entire range of
wavelengths. This is referred to as the
pigment’s absorption spectrum. Because each pigment’s absorption spectrum is
unique, like a fingerprint, it can be used to identify the pigment.

During lab, you will use TLC to separate a mixture of pigments that have
been extracted from spinach leaf cells. After you have separated the pigments, you will use a scanning spectrophotometer to plot the
absorption spectra for 3 of the pigments and then use this information to identify
the 3 pigments.

Your Turn

In cell and molecular biology, what are
electrophoresis and chromatography used for?